tbuckley/cabot-search huggingface dataset:
Abstract P1084: Cardiomyocyte-specific Overexpression Of Ccnd2 Via Modified Mrnafor Remuscularization In Hearts With Myocardial Infarction
Authors: Jiacheng Sun, Lu Wang, Rachel Matthews, Gregory Walcott, YU-AN LU, Yuhua Wei, Yang Zhou, Lior Zangi, Jianyi J Zhang
Journal: Circulation Research (2023)

Introduction: The adult mammalian heart has limited regenerative capacity due to postnatal cardiomyocyte (CM) cell cyclearrest. With the success of modified mRNA (modRNA) vaccine against SARS-CoV-2, modRNA technology offers efficient, transient, safe, nonimmunogenic, and controlled mRNA delivery without any risk of genomic integration. Using our previously reported cardiomyocyte specific modRNA translation system (CM SMRTs), we investigated the myocardial regenerative potential and therapeutic outcome of transient and specific cardiomyocyte overexpression of cyclin D2 (CCND2) in a pig model of myocardial infarction (MI). Methods: Modified mRNA was synthesized by in vitro transcription. AMI was induced by occlusion of the left anterior descending coronary artery for 60 minutes with reperfusion. Pigs were randomly assigned to vehicle group (1ml sucrose-citrate buffer, n=6), nGFP-CM SMRTs group (7.5mg CM-specific nuclear GFP modRNA, n=7)and CCND2-CM SMRTs group (7.5mg CM-specific CCND2 modRNA, n=8). CM cell cycle activity was measured 3 days after injection by immunostaining for proliferation marker Ki67, M-phase marker phospho-histone H3 (PH3) and cytokinetic marker aurora B kinase (AuB). Cardiac function was evaluated by echo and cardiac MRI (cMRI) on day 28; infarct size was determined via late gadolinium enhancement cMRI; arrhythmia incidence was assessed via implanted loop recorders. Results: The short-term in vivo study demonstrated a highly specific overexpression of targeted genes in CMs. CCND2-CM SMRTs significantly promoted CM cell cycle activation that was evidenced by significant increase of cell cycle markers compared with nGFP-CM SMRTs group: Ki67 (4.21±0.34% vs. 0.81±0.05%), PH3 (0.51±0.06% vs. 0.13±0.03%), and Symmetric AuB (0.035±0.007/mm 2 vs. 0.000±0.000/mm 2 ), 3 days after AMI and modRNA treatment. The long-term follow-up of the animals also showed improved heart function (EF: 50.34±1.94% vs. 38.60±1.18%) and decreased scar size (8.80±1.07% vs. 22.77±2.77%). The frequency of arrhythmic events in animals between all groups were similar. Conclusion: The results in large animal model of AMI demonstrate remuscularization and therapeutic potential of cardiomyocyte-specific overexpression of CCND2 via modRNA.
Abstract P3031: The Cross Talk Between Increased Abdominal Adiposity And Calcification Of Aortic Valve
Authors: Rihab Bouchareb
Journal: Circulation Research (2023)

Calcified aortic valve disease (CAVD) is the most prevalent valvular heart disease in western populations affecting nearly 2.5% of individuals over 65. Currently, there is no viable pharmacological treatment to stop the disease's progression or activate mineral regression. The only effective therapy to treat CAVD is surgical valve replacement or trans-catheterization. Obesity is the major common nutritional disorder worldwide and is one of the important risk factors for CAVD. It has been shown that DLK1-/- mice have increased abdominal adiposity. Indeed Dlk1 plays an essential role in regulating adipocytes differentiation and proliferation. Recently, it has been shown that there is an association between fat tissue accumulation and an increased risk of aortic stenosis. We hypothesize that dysregulation of DLK1 stimulates adiposity, increases circulating lipids, enhances valve lipids retention, and stimulates calcification. Herein, our objective is to examine the expression of DLK1 in calcified aortic valves and its impact on lipid retention. Method: We examined the effect of Dlk1 loss of function on lipid retention in aortic valve tissue in ApoE-/- Dlk1-/- mice fed with a high-fat diet and in human explanted aortic valves using immuno-staining. Results: The complete deletion of DLK-1 in ApoE-/- mice were associated with an increased oxidized-LDL deposit in aortic valve tissue. In addition, circulating cholesterol (420 ± 8 mg/dl vs 250± 12 mg/dl, P=0.002) and triglyceride concentration (200 ± 3 mg/dl vs 50± 10 mg/dl, P&lt;0.001) were higher in ApoE-/- DLK1-/- compared to apoE-/- mice. An increased valvular calcification accompanied these changes, as shown by the over-expression of RUNX2 and the increased aortic jet velocity in ApoE-/- DLK1-/- mice. Interestingly the overexpression of DLK1 in isolated human valve cells decreased calcification and lipid retention in vitro. Conclusion: This study showed the significant role of Dlk1-derived abdominal adiposity in valve-lipid retention and calcification.
Abstract P2057: The Role Of Cardiac Neurofibromin-2 In Chronic Pressure Overload
Authors: Satvik Mareedu, Yu Zhang, Wataru Mizushima, Jamie Francisco, Jin Guan, Zhi Yang, Shinichi Oka, Peiyong Zhai, Maha Abdellatif, Dominic Del Re
Journal: Circulation Research (2023)

Department of Cell Biology and Molecular Medicine, New Jersey Medical School, Rutgers University, Newark, NJ 07103.Neurofibromin 2 (NF2) is a tumor suppressor that can engage the Hippo signaling pathway and modulate cell proliferation and survival. We previously demonstrated that NF2 mediates cardiomyocyte apoptosis and injury caused by acute myocardial infarction. However, the function of NF2 in the heart remains largely uncharacterized. Our current study sought to determine whether NF2 modulates heart failure due to chronic stress. We used a transverse aortic constriction (TAC) model in WT C57BL/6J mice to generate chronic pressure overload (PO) stress, which elicits cardiac remodeling and failure. We found that NF2 is transiently upregulated in mouse myocardium in response to early phase of PO, and is downregulated during late phase PO. We generated cardiomyocyte-specific NF2 knockout (cKO) mice, which had normal cardiac morphology and function at baseline. Following TAC, NF2 cKO hearts unexpectedly showed worsened cardiac function, assessed by echocardiography and hemodynamic analysis, compared to controls. RNAseq analysis indicated downregulation of several metabolic pathways including oxidative phosphorylation and fatty acid oxidation in NF2 cKO hearts at baseline. Additionally, we observed reduced ATP content in baseline NF2 cKO hearts compared to controls. Fractionation experiments indicated that nuclear NF2 is enriched following PO stress. RNAseq revealed the downregulation of ERRβ and ERRγ in NF2 cKO hearts, which was confirmed by qPCR. Experiments employing neonatal rat ventricular myocytes (NRMVs) confirmed that NF2 regulated expression of ERRβ and ERRγ, and DNA pulldown assays demonstrated NF2 association with ERRβ and ERRγ proximal promoters. Luciferase reporter assays demonstrated that NF2 modulates ERR function in NRVMs. Based on these findings, we propose that transient upregulation of myocardial NF2 expression during PO stress is compensatory and regulates metabolic gene expression according to energy demand.
Abstract P2184: Isocitrate Dehydrogenase 1 Regulates Cardiac Metabolic Adaptation During Oncometabolic Stress
Authors: kyoungmin Kim, Yaqi Gao, Brandon Faubert, Nathaniel Snyder, Ralph DeBerardinis, Anja Karlstaedt
Journal: Circulation Research (2023)

Cardiovascular disease and cancer are the two leading causes of morbidity and mortality in the world. Metabolic dysregulation of cancer cells extends beyond the tumor microenvironment and increases risk for cardiovascular diseases. One common somatic mutation in cancer cells affects isocitrate dehydrogenase (IDH) 1 and 2, which catalyzes the oxidative decarboxylation of isocitrate to alpha-ketoglutarate in the cytosol and mitochondria, respectively. IDH1 and 2 mutations cause the production of the oncometabolite D-2-hydroxyglutarate (D2-HG) which allosterically inhibits alpha-ketoglutarate dehydrogenase (alpha-KGDH) and is associated with reduced cardiac contractile function. We combined stable isotope tracer studies with computational modeling to study the fundamental role of IDH isoforms for cardiac adaptation during oncometabolic stress. We uncovered an unexpected cardiac phenotype that expands the role of IDH1 in the heart beyond oxidative metabolism. Using mass spectrometry-based metabolomics, we quantified the stable isotopomer distributions from glucose and glutamine in perfused working rat hearts and isolated adult ventricular cardiomyocytes. Our analysis revealed that defective mitochondrial metabolism causes redirection of carbon flux oxidative towards reductive pathways. Reductive carboxylation of alpha-KGDH increases glutamine uptake and glutamine-derived citrate formation in both working rat heart perfusions and cultured adult mouse ventricular cardiomyocytes. To identify which IDH isoform is responsible for redirecting carbon flux, we developed knockout models of IDH1, 2, and 3 in adult mouse ventricular cardiomyocytes. Loss of IDH1 expression impaired reductive formation of citrate and caused functional defects in cardiomyocytes. Lastly, epigenetic analyses of histone marks revealed that IDH1 causes widespread alterations in histone acetylation and tri-methylation. Our results highlight a novel role for IDH1 in cardiac metabolism and transcriptional control of metabolic adaptation in response to tumor-mediated stress and provide evidence of how reductive-citrate formation may induce epigenetic modifications in the heart.
Abstract P2056: The Transcription Factor TFEB Protects Against Cardiac Hypertrophy By Increasing Lysosomal Capacity And Activating AMPKα
Authors: Subhajit Das Gupta, Daniel Daou, Herman May, Nan Jiang, Nicholas Nguyen, Anip Anand, Camila Iansen Irion, Valeria Garrido-Moreno, Francisco Olivares-Silva, Abhinav Diwan, Thomas G Gillette, Joseph A Hill
Journal: Circulation Research (2023)

Introduction: Pathological cardiac remodeling in response to elevated afterload can eventually progress to heart failure. Lysosome-assisted processes such as autophagy play an important role in this remodeling. The lysosome is also a nexus for some of the same signaling pathways that are involved in hypertrophic growth. The transcription factor EB (TFEB) has emerged as a key regulator of lysosomal genes in multiple tissues, especially in response to external stress. Methods and Results: Here, we report that TFEB is rapidly activated and translocates to the nucleus in cardiomyocytes under hypertrophic stress conditions, activating a lysosomal gene program independent of autophagy gene changes. We engineered a cardiomyocyte-specific TFEB knockout mouse (CTKO). At baseline, contractile function measured by echocardiography appears normal in these mice compared with their Cre-negative littermates. However, in the setting of pressure-overload stress induced by TAC (thoracic aortic constriction), the CTKO mice manifest an exacerbated hypertrophic response leading quickly to heart failure. Unlike the WT hearts, CTKO hearts fail to increase lysosomal capacity after TAC and manifest an increase in steady-state levels of autophagosome-associated proteins such as LC3II and p62, as well as accumulation of ubiquitinated proteins, suggesting a decrease in protein turnover. Surprisingly, AMPK signaling, while normal at baseline is significantly decreased in the CTKO hearts after TAC. This reliance on TFEB for PE-induced AMPK signaling is also observed in vitro , as are the anti-hypertrophic effects of TFEB activation, supporting a direct role of TFEB in this process. Finally, we show that exogenous activation of AMPK in the absence of TFEB can completely rescue the exacerbated hypertrophic response both in vitro and in vivo , independent of lysosomal function. Surprisingly, blunting of the hypertrophic response had no impact on the decreased cardiac function observed in the TAC-treated CTKO mice. Conclusion: Overall, our findings suggest that TFEB antagonizes pathological hypertrophic cardiac remodeling through up-regulation of lysosomal capacity and AMPK signaling.
Abstract P2052: Fibroblast-specific TGF-b Signaling Protects The Infarcted Myocardium From Adipose Tissue Infiltration
Authors: Izabela Tuleta, Harikrishnan Venugopal, Claudio Humeres, Silvia C Hernandez, Akihiko Kubota, Anis Hanna, Nikolaos G Frangogiannis
Journal: Circulation Research (2023)

Background and Objectives: Repair of the infarcted heart requires timely phenotypic transitions of fibroblasts, the central effector cells involved in scar formation and matrix remodeling. TGF-b superfamily members are critical regulators of fibroblast phenotype, acting through TGF-b receptors (TbR) and downstream activation of Smad-dependent and Smad-independent cascades. We have previously demonstrated that fibroblast-specific Smad3 plays a central role in repair of the infarcted heart; however, the role of TGF-b/TbR signaling in regulation of infarct fibroblast transitions has not been investigated. In the current study we examine the role of fibroblast-specific TGF-b signaling in repair of the infarcted heart, using mice with inducible fibroblast-specific disruption of TbRII (FTbRIIKO mice), the only type 2 receptor mediating the effects of all 3 TGB-b isoforms. Findings: FTbRIIKO animals exhibited increased mortality after myocardial infarction (MI), caused by cardiac rupture. In comparison with TbRII fl/fl mice, TbRII KO mice had significant thinning of the anterior wall 28 days after MI, without significant differences in ventricular function. Fibroblast-specific TGF-b signaling loss did not affect myofibroblast infiltration 7 days after MI. 28 days after MI, brightfield and polarized microscopy showed that, although scar size was comparable between the groups, collagen content and the width of collagen fibers were reduced within infarcted zones of FTbRIIKO vs. FTbRII fl/fl animals. Moreover, FTbRIIKO mice exhibited a striking deposition of adipocytes, occupying up to 40% of the infarct zones (fatty infiltration: FTbRIIKO, 28.7±6.2% vs floxed 0±0%, p&lt;0.001, n=6-8). Infarct fibroblasts harvested from FTbRIIKO mice 7 days after MI had increased expression of adipocyte genes, such as perilipin, leptin receptor, and PPARgamma. Conclusions: Fibroblast-specific TGF-β signaling protects the infarcted heart from cardiac rupture and prevents fatty infiltration of the infarct. These effects may reflect critical actions of TGF-bs on fibroblast phenotypic transitions and inhibition of fibroblast to adipocyte conversion.
Abstract P3142: Engineering A Genetically Encoded Enhancer Of L-type Ca <sup>2+</sup> Currents
Authors: Pedro Javier J Del Rivero Morfin, Srinidhi Jayaraman, Lin Yang, Audrey Kochiss, Henry M Colecraft, Steven Marx, Manu Ben-Johny
Journal: Circulation Research (2023)

L-type calcium channels (LTCC) play a crucial role as they serve as vital portals for Ca 2+ entry into cardiac myocytes, neurons, and other physiological systems. Altered LTCC function is linked to various pathologies including cardiac arrhythmias and neurodevelopmental disorders. In particular, genetic studies have identified loss-of-function mutations in Ca V 1.2 channels in patients with shortened QT interval and Brugada Syndrome, as well as neuropsychiatric disorders. Given the broad physiological importance of LTCC in various tissues, conventional pharmacological agonists of LTCC may yield off-target effects and toxicity. Therefore, new approaches to upregulate LTCC function in a cell-type dependent manner is sought after. To do so, we here develop a genetically encoded enhancer of calcium channels (GeeCC, “geek”) by leveraging both a cardiac specific L-type channel modulator, leucine rich repeat containing protein 10 (Lrrc10), and selective nanobodies that target the LTCC complex. Specifically, Lrrc10 co-expression markedly upregulates LTCC currents. Furthermore, mutations in Lrrc10 are linked to cardiac diseases, including cardiomyopathies. Through extensive structure-function analysis, we identified a minimal action domain within Lrrc10 that is critical for functional modulation of LTCC. However, as this segment alone has a low affinity for the channel complex, its expression alone yielded minimal changes in Ca 2+ current amplitude. However, fusion of this Lrrc10 domain to a nanobody (nb.F3) that targets the Ca 2+ channel beta subunit resulted in a significant increase in the LTCC current amplitude. In depth analysis revealed exquisite selectivity of GeeCC for Ca V 1.2/Ca V 1.3 channels. In mouse cardiomyocytes, deployment of GeeCC via adenovirus enhances endogenous L-type currents when compared to uninfected controls or cardiomyocytes infected with nanobodies alone. Additionally, in neurons, co-expression of GeeCC enhanced excitation-transcription coupling. In all, we have developed a genetically-encoded approach to selectively enhance L-type currents. This strategy may enable new insights into cardiovascular and neuronal physiology and pathophysiology, and may have potential therapeutic applications.
Abstract P2174: Transcriptomic Profiling Of Immune Checkpoint Inhibitor Myocarditis Reveals Novel Therapeutic Targets
Authors: Juan Qin, Shaoyi Zhang, Amir Munir, Yen-Chou Chen, Kuan Zhang, Jiaqi Li, Joe-Elie Salem, Javid Moslehi
Journal: Circulation Research (2023)

Introduction: Immune checkpoint inhibitor (ICI) therapy has revolutionized the treatment of advanced cancers. However, the increased use of ICI has also resulted in a higher incidence of ICI-related cardiovascular side effects, including myocarditis, which has a high mortality rate of nearly 50%. Understanding the mechanisms underlying ICI-related cardiotoxicity is essential for identifying new therapeutic targets and improving patient survival. Hypothesis: We hypothesized that ICI-myocarditis has a unique transcriptomic profile that could be used to identify new drug targets. Methods: To test our hypothesis, we isolated RNA from our ICI-myocarditis mouse hearts and performed RNA-seq using the Illumina NovaSeq 6000. We also reanalyzed a human IC-myocarditis dataset downloaded from the Gene Expression Omnibus database. Results: Our results demonstrated that in both our animal models and patients, the upregulated pathways in ICI myocarditis heart tissues were enriched for regulation of T cell activation, leukocyte adhesion, and multiple cytokine signaling pathways. Conversely, downregulated pathways included those involved in cellular respiration, oxidative phosphorylation, and cardiac conduction. The most significantly upregulated genes were involved in innate and adaptive immune responses, including the CD8-dependent gene program and response to interferon-gamma of macrophages. By querying the Drug Gene Interaction Database, we identified several novel biologically plausible therapeutic targets. Correspondingly, a number of FDA-approved drugs were revealed, including monoclonal or chimeric antibodies such as Tocilizumab and Alemtuzumab, recombinant proteins like Abatacept and Anakinra, as well as small molecules such as Ruxolitinib and Imiquimod. Conclusions: Our study is the first to use the entire transcriptomic profile of ICI myocarditis in both preclinical models and patients for drug discovery. Our results demonstrate that ICI myocarditis has a distinct gene expression profile compared to healthy and other cardiac diseases. By querying a drug-gene interaction database, we identified several novel drug targets and possible treatments for ICI-related cardiotoxicity.
Abstract P2062: Distinct Stiffness-induced Mechanical Memory Induction And Maintenance In Cardiomyocytes And Cardiac Fibroblasts
Authors: NESRINE BOUHRIRA, Alexia Vite, Kenneth B Margulies
Journal: Circulation Research (2023)

Recognizing that cells “feel” and respond to their mechanical environment, recent studies demonstrate that many cells exhibit a phenomenon of “mechanical memory” in which features induced by prior mechanical cues persist after the mechanical stimulus has ceased. While there is a general recognition that different cell types exhibit different responses to changes in extracellular matrix stiffening, the phenomenon of mechanical memory within myocardial cell types has received little attention to date. To probe the dynamics of mechanical memory in cardiomyocytes(CMs) and cardiac fibroblasts (CFs), we developed a magnetorheological elastomer (MRE) cell culture substrate with tunable and reversible stiffness spanning the range from normal to diseased myocardium. Employing IPSC-derived CMs, and CFs while measuring canonical responses to matrix stiffening: cell area, myofibroblasts activation, and chromatin condensation parameter reveals distinct temporal dynamics of mechanical memory induction in each cell type where mechanical memory is invoked much earlier in CMs than in CFs. Experiments using specific inhibitors of cytoskeletal components revealed that induction of mechanical memory is actin-dependent in CFs and microtubule-dependent in CMs. Prevention of α-tubulin detyrosination blocks both induction and maintenance of mechanical memory in CMs. Overall, these results emphasize the distinct temporal dynamics of mechanical memory in CMs and CFs with different cytoskeletal mediators responsible for inducing and maintaining the stiffness activated phenotype. Due to its flexibility, this model is broadly applicable to future studies interrogating mechanotransduction and mechanical memory in the heart and might inform strategies for attenuating the impact of load-dependent pathology and excess myocardial stiffness.
Abstract P3151: A Unique Profile Of Extracellular Vesicles In Marfan Patients With Thoracic Aortic Aneurysm
Authors: J. Nathaniel Diehl, Amelia Ray, Lauren B Collins, Andrew Peterson, Kyle Alexander, Jacob Boutros, John S Ikonomidis, Adam W Akerman
Journal: Circulation Research (2023)

Objectives: Extracellular vesicles (EVs) [exosomes and microvesicles] have significant diagnostic and therapeutic potential, but clinical application is limited by disparate methods of data collection. Exosomes are 30-100nm, and microvesicles are 100-300nm lipid bilayer vesicles that originate from all cell types. Marfan Syndrome (MFS) is a connective tissue disorder. Thoracic aortic aneurysm (TAA) formation and dissection are the leading causes of death in MFS patients. Early identification of TAAs in MFS patients is imperative and will mitigate morbidity and mortality. This study’s goal was to develop a reproducible workflow for isolation and quantification of EVs 30-300nm and test the hypothesis that number and size distribution are altered in MFS patients with TAA. Methods: A method of standardized reproducible measurement of EVs isolated from plasma was developed. This includes isolation by automated size exclusion chromatography and quantification of size distribution by tunable resistive pulse sensing. Characteristic markers specific to both EV subtypes were confirmed by immunoblot. Results: EVs isolated from control plasma (n=12) and Marfan TAA plasma (n=6) were measured for concentration (control 0.60 x 10 11 particles/mL, MFS TAA 1.47 x 10 11 particles/mL), mean diameter (control 77.75nm, MFS TAA 69.92nm), mode diameter (control 65.50nm, MFS TAA 60.50nm), and d90/d10 ratio (control 1.68, MFS TAA 271 1.57). Conclusion: MFS patients with TAA have more EVs with a smaller diameter, suggesting increased export and a shift in ratio of microvesicles:exosomes. This may be used as a diagnostic tool or potential therapeutic target. This scalable protocol ensures recovery of intact EVs suitable for downstream biochemical and functional analyses. Simultaneous measurement quantifies EV concentration and size distribution absolutely, correcting for variations, providing a novel method of standardization.